Cat. No.: IAPRCA-50 (for 50T)

Description

PrimeIAmp™ Primase Rolling Circle Amplification (RCA) Kit (50T) utilizes the Primase Pol enzyme to generate amplification primers, employing a similar mechanism to the Tth PrimPol Primase used in the TruePrime RCA Kit. This amplification method, which does not rely on Random Hexamers, ensures precise amplification of the template and avoids the introduction of uncertain bases from primers. Primase Pol binds to ssDNA and synthesizes short complementary primers using dNTPs as substrates. Subsequently, under the action of the highly faithful and robust phi29 HT DNA Polymerase, ssDNA is continuously synthesized. The newly synthesized ssDNA can serve as a substrate for Primase Pol to re-synthesize complementary primers, thus continuing the amplification process and forming dsDNA.

The kit is equipped with dsDNA denaturation buffer (Buffer D), allowing both ssDNA and dsDNA to effectively serve as amplification substrates. With 1ng of circular plasmid template, 3μg of amplification product can be obtained within 2 hours. Using 10ng of genomic DNA (100-200kb) as a template, 1-2μg of product can be obtained. The final amplification product is a high-molecular-weight network-like dsDNA, which can be used for enzymatic digestion, sequencing, and ligation.

Content

• 10×phi29 Buffer: 150 μl
• Buffer D: 150 μl
• Buffer N: 150 μl
• phi29 HT DNA Polymerase (10U/µl): 50 μl
• Primase Pol: 50 μl
• 10mM dNTP: 75 μl
• RNase Free H2O: 1 ml

Features

• Precise Amplification: Ensures precise amplification of the template by utilizing Primase Pol enzyme, which minimizes the introduction of uncertain bases from primers.
• Efficient Primer Generation: The Primase Pol enzyme synthesizes short complementary primers on ssDNA using dNTPs as substrates, facilitating efficient primer generation.
• High Fidelity: Amplification facilitated by phi29 HT DNA polymerase, known for its high fidelity and robust synthesis capabilities, ensures reliable results.
• Continuous Amplification: Continuous synthesis of ssDNA allows for continuous primer synthesis and amplification, enhancing efficiency.
• Versatile Substrate Compatibility: The kit includes dsDNA denaturation buffer (Buffer D), enabling both ssDNA and dsDNA to serve as effective amplification substrates, offering versatility in experimental design.
• High Yield: Offers high amplification yield, with 1ng of circular plasmid template yielding 3μg of amplification product within 2 hours, and 10ng of genomic DNA (100-200kb) yielding 1-2μg of product.
• Versatile Applications: The final amplification product is high-molecular-weight network-like dsDNA, suitable for various downstream applications including enzymatic digestion, sequencing, and ligation.

Storage

The minimum shelf life is 2 years at -20°C. Avoid repeated freezing and thawing.

Instruction: Protocol

Learn more: Choosing Your Best Phi29 DNA Polymerase and Padlock Primers to Start Your Rolling Circle Amplification

Related: 

• phi29 DNA Polymerase
• phi29 DNA Polymerase (glycerol-free)
• phi29 DNA Polymerase (lyophilized)
• phi29 HT DNA Polymerase
• PrimeIAmp™ Rolling Circle Amplification Kit
• PrimeIAmp™ Primase Rolling Circle Amplification (RCA) Kit
• Tth PrimPol Primase

Only for research and not intended for treatment of humans or animals

SBS Genetech is a long-term sponsor of Cold Spring Harbor Laboratory